Ferritin Plays A Role In The Pathogenesis Of Adult-onset Still’s Disease By Facilitating Neutrophils Extracellular Traps

Objective:Adult-onset Still’s disease(AOSD)is a complicated systemic inflammatory disease.It’s characterized by neutrophilia and high levels of serum ferritin.The release of neutrophil extracellular traps(NETs)has been demonstrated as an effector mechanism of neutrophils in many inflammatory diseases.We investigated whether aberrant NETs occur in AOSD,examined the pro-inflammatory properties of NETs and explored the potential capacity of ferritin to trigger NETs.Methods:Sera from 73 AOSD patients,30 rheumatoid arthritis,30 systemic lupus erythemosus and 40 healthy controls were collected to determine the level of cell-free DNA,and NET-DNA complexes,including citrullinated histone 3(cit H3)-DNA,neutrophils elastase(NE)-DNA and myeloperoxidase(MPO)-DNA complexes.NET formation was analyzed using immunofluorescence(IF)and Pico Green.NADPH oxidase-derived reactive oxygen species(ROS)and mitochondrial ROS production were determined by flow cytometry and IF.NLRP3 inflammasome activation in THP-1 cells and CD14~+monocytes,and pro-inflammatory macrophages activation stimulated with NET-DNA were determined.The copy number of mitochondrial DNA in NETs and plasma was detected by RT-PCR.Results:The levels of cell-free DNA and NET-DNA complexes were significantly higher in the serum of patients with AOSD,and neutrophils from AOSD patients were prone to spontaneously release more NETs,an effect that can be replicated by treating healthy control neutrophils with ferritin.Moreover,ferritin induced-NET release was in a ROS-dependent manner.DNA purified from AOSD NETs activated NLRP3 inflammasome,accelerated the activation of CD68~+CD86~+macrophages,and faciliated the expression level of IL-1β,IL-6 and TNF-α.Finally,higher copy number of mt DNA were detected in NETs and plasma from AOSD patients.Conclusions:Our findings indicate that enhanced NET formation plays a pathogenetic role in AOSD though stimulation of NLRP3 and pro-inflammatory macrophages.In addition,we found a new pathway of ferritin-NETs-inflammatory factors.It provides theoretical and experimental basis for developing more effective diagnostic and therapeutic methods of AOSD.